The Differentiation Of Mouse Embryonic Neural Stem Cells And Otocyst Cells In Vitro

Objective:To find the potency of neural stem cells and otocyst cells of differentiating into inner ear hair cells and to value the relationship of the neural stem cells with the regeneration of the hair cells.Methods:Neural tubes of hindbrain and otocysts were dissected out from10day’s embryos, and then were dissociated in Hank’s Buffered Salt Solution (HBSS) separately. After being digested, the cells were cultured in serum free medium with bFGF and EGF in vitro. When the neurospheres or otospheres were found, cells were purified with neurosphere assay. The immunocytochemistry (ICC) and RT-PCR were taken to see what neurospheres and otospheres were composed of. These cells were cultured in serum medium for15days and then detected with ICC with the antibodies against nestin and myosinVIIa as the markers of stem cells and hair cells. RT-PCR was also performed to detect the expression of transcript factors Pax6, SOX2, BMP7, Jagged-1, p27Kipl, MyosinⅦa and Gfil.Results:The neural tubes and otocysts were obtained successfully from the embryos. The neurospheres and otospheres were fromed when cultured in serum free medium in vitro. The nestin was expressed in cells from neurospheres and otospheres. After15days’culture in the medium containing serum, we found both RNA and proteins of MyosinⅦa in cells derived from neurospheres and otospheres.Conclution:The Stem cells from neural tubes and otocysts have the ability to differentiate into hair cell-like cells in vitro. Neural stem cells and otocyst cells will be a fine tool to research the gene expression and regulation in the development of the inner ear.